The proposed research will advance preclinical characterization and validation of a novel drug for asthma. A growing body of research has uncovered functional gamma amino butyric acid type A receptor (GABAAR) signaling in non-neuronal cells. Our research has demonstrated that ligands activating GABAAR subtypes located on airway smooth muscle and immune cells reduce airway hyper-responsiveness (AHR) and inflammation when orally administered in animal asthma models. The significance of this innovation is a novel receptor targeting and drug design strategy for a safe and effective oral asthma treatment. Innovative new treatments are needed to cost-effectively control symptoms, reduce adverse effects, improve compliance by avoiding inhaler use, and provide better solutions for steroid-resistant asthma patients. The long-term goal of this research is approval of a first-in-class oral drug for asthma. The objective of the Phase I STTR contract is optimization of lead compound MIDD0301 via a formulation that enhances oral bioavailability as determined by drug pharmacokinetics (PK), metabolism, and clearance. To establish compound safety, a battery of safety pharmacology testing will be conducted using approaches set forth in regulatory guidelines. Our central hypothesis based on preliminary data is that therapeutic levels of MIDD0301 are readily achieved in the lung and GABAARs can be targeted safely with anionic ligands for asthma treatment. The rationale for these nonclinical studies is the need to identify a formulation of MIDD0301 suitable for clinical testing and corresponding drug exposure and safety verification to estimate an initial safe starting dose and dose range for future human trials. Our hypothesis will be tested within three Specific Aims: (1) Identify the most bioavailable polymorphic form of MIDD0301 and its salts; (2) establish MIDD0301 safety by safety pharmacology testing; and (3) identify and investigate possible MIDD0301 metabolites. The significance of this research is advancement of safe and effective drug candidate that controls asthma symptoms, avoids steroid resistance, and improves compliance. Furthermore, it is expected that development of MIDD0301 will lay the foundation for future drug research that targets peripheral GABAARs for other immune-inflammatory diseases in addition to asthma. The innovation of this research is the validation of a new therapeutic target for asthma common to airway smooth muscle and immune cells that can be modulated with a single small molecule to control airway hyperresponsiveness and inflammation, two pathophysiological hallmarks of asthma. This research will expand crosscutting knowledge of a unified paracrine GABAergic system in the lung that can be applied to other lung disorders. .